Characterization of the heat shock response in Escherichia coli has allowed identification of at least twenty heat shock proteins (hsps), some of which are essential for viability at normal temperatures. About half of the hsps in E. coli have been characterized. In spite of rapid progress on the biochemistry of the major hsps, much is still to be learned about their physiological roles. This proposal describes a systematic examination of the effect of heat shock on cell envelope proteins in E. coli employing cell fractionation and electrophoretic analysis. The two specific aims are to determine why the major hsps DnaJ and DnaK fractionate with the membrane, and to identify membrane and periplasmic proteins that are induced by heat shock. Experiments will determine if the membrane association observed for DnaJ and DnaK is artifactual, if they are peripheral membrane proteins or whether they associate with the membrane at a specific point in the cell cycle. When membrane hsps are identified, their inclusion in the heat shock regulon will be determined by their dependence on sigma 32. Future work will test whether membrane hsps bind to lipids, can be chromatographed from membrane extracts using affinity chromatography with immobilized hsps, and respond to other stresses, including low pH, carbon starvation and antibiotic treatment. Since hsps in the membrane can be expected to play roles in cell division, protein translocation and perhaps stabilization of the cell wall, this study may locate such proteins which are minor constituents that have eluded identification so far, thus contributing to a general understanding of the role of the membrane in these processes. Studies of E. coli hsps provide models for numerous pathogenic bacteria found to contain hsps that cross-react to the major hsps of E. coli; several human pathogens have hsps on their cell surfaces. These hsps are highly immunogenic: in pathogens such as Borrelia burgdorferi (lyme disease) and Mycobacterium tuberculosis (adjuvant arthritis in rats) the hsps are the dominant antigens in the host.